Immunoreactive Calcitonin Levels Research Articles

Validation, role in perioperative assessment, and clinical applications of an immunoradiometric assay for human calcitonin

We have evaluated a new immunoradiometric assay (IRMA) for calcitonin (CT) (Medgenix, Belgium). The detection limit of immunoreactive CT (IR-CT) was 2.8 pg/ml, and plasma levels of IR-CT of 134 healthy adults (dynamic range, 2.8–1000 pg/ml) were < 19 pg/ml (median 4.1 pg/ml). Both basal and pentagastrin-stimulated IR-CT levels were <30 pg/ml and were well separated from the levels in patients with medullary thyroid carcinoma (MTC) (46,000–29,000 pg/ml, n = 38). In preoperative plasma samples in five patients with C-cell hyperplasia, CT IRMA confirmed the diagnosis, although in two patients RIA failed to detect a significant increase of IR-CT. By modifying the IRMA, we could identify residual tumor tissues perioperatively in patients undergoing thyroidectomy for MTC. RIA of gel-permeation chromatography showed four IR-CT peaks, but IRMA had only three peaks, suggesting that IRMA is more specific. CT-IRMA is more specific, sensitive, robust, rapid, and reliable than RIA. In addition, neat plasma can be used directly in a single step in CT-IRMA.

Immunoreactive "calcitonin-like" material in heroin addicts: varying reactivity with different antibodies.

High levels of immunoreactive calcitonin (iCT) in the blood of heroin addicts were previously reported. As it is well known that multiple forms of calcitonin exist in the blood and in tissues, the purpose of the present study was to investigate the immunological nature of the CT-like immunoreactive material found in the blood of these subjects. We investigated 25 addicts, who had been using heroin for more than one year and were hospitalized for a 2 week detoxication program. Blood samples were drawn at the start of the program (when the subjects were still on heroin) and after 5 and 12 days of abstinence from heroin. Twenty-five healthy subjects served as controls. We used 2 commercial RIA kits, calibrated against the same reference material (WHO 70-234), but employing different antisera. One antiserum substantially confirmed the previous findings of increased levels of calcitonin during heroin use, but the other seemed to exclude any change in the hormone concentrations. This suggests that the "calcitonin like" material found in heroin addicts contains some epitopes similar to those found in the calcitonin standard which are detected by the first antiserum. However it lacks other epitopes which are also present in calcitonin standard and which are recognized by the second antiserum. Therefore, this substance seems to be different from the standard human calcitonin 1-32. A possible involvement of a calcitonin analogue (precursor or metabolite) in the biochemical changes occurring during chronic opiate use is suggested.

The effect of hypermagnesaemia on serum immunoreactive calcitonin levels in normal human subjects.

The effect of hypermagnesaemia on serum levels of immunoreactive calcitonin was studied in normal human subjects. After iv administration of magnesium sulphate over 120 min, the mean (+/- SEM) serum magnesium concentration rose from the baseline level of 0.9 +/- 0.1 to 2.6 +/- 0.3 mmol/l (P less than 0.01), and thereafter remained higher than the baseline level. The magnesium infusion caused a significant increase in serum immunoreactive calcitonin levels (P less than 0.01). The rise in serum magnesium concentration was accompanied by a significant decrease in the concentrations of corrected serum calcium and whole blood ionized calcium (P less than 0.01, P less than 0.01 respectively). Our results suggest that hypermagnesaemia causes an increase in serum immunoreactive calcitonin levels in normal human subjects despite a decrease in the concentrations of corrected serum calcium and whole blood ionized calcium.

Calcitonin concentrations in lung cancer and non-malignant pulmonary diseases.

The serum immunoreactive calcitonin (ICT) concentration was estimated in 107 patients with lung cancer of various cell types, 46 normal subjects and 24 non-cancer patients with various pulmonary diseases. Eighty-one of the cancer patients had localized lung cancer and the rest 26 primary lung cancer with metastases to other organs. The ICT was found significantly increased in the lung cancer patients. The highest ICT values were found in patients with lung cancer and metastases to other organs, especially to the liver. In localized disease, patients with small-cell carcinoma of the lung showed the highest, while those with adenocarcinoma and squamous-cell type demonstrated the lowest ICT concentrations. In 46 patients with lung cancer, serial determinations of serum ICT levels showed significantly decreased calcitonin levels in the patients who responded well to therapy with cytostatics, X-rays or both after more than 2 months of therapy. Fluctuations of ICT levels were in agreement with the regression or progression of the disease.

Immunoextracted calcitonin in milk and plasma from totally thyroidectomized women. Evidence of monomeric calcitonin in plasma during pregnancy and lactation.

The level of immunoreactive calcitonin in the first produced breast milk was in totally thyroidectomized (TX) women 713 +/- 307 pg-eq/ml (mean +/- SD, N = 7) and in control women 772 +/- 329 pg-eq/ml (N = 33), i.e. about 45 times higher than in plasma (see below). On gel chromatography of immunoextracted milk from TX women, immunoreactive calcitonin appeared as high molecular weight forms in the same pattern as in milk from healthy women when the same antiserum was used for immunoextraction and radioimmunoassay (RIA). In another series of experiments, a new antiserum raised in rabbits was used for measurement of immunoreactive calcitonin after immunoextraction with 1. Plasma levels of immunoreactive calcitonin in the TX women during pregnancy were 16 +/- 6 pg-eq/ml (N = 6) and during lactation 14 +/- 7 pg-eq/ml (N = 5). Immunoreactive calcitonin was undetectable (less than 8 pg/ml) in plasma from those TX women in whom lactation had stopped (N = 5). Immunoextraction and gel chromatography of plasma collected during pregnancy and lactation from the TX women showed that the immunoreactive calcitonin present eluted in the region of monomeric calcitonin with both antiserum 1 and 2. In conclusion, high concentrations of high molecular weight forms of of immunoreactive calcitonin have been demonstrated in milk from TX patients, most probably devoid of any calcitonin-producing thyroid C-cells. This points to a local production site in the mammary gland.(ABSTRACT TRUNCATED AT 250 WORDS)

Immunoextracted calcitonin from human seminal plasma constitutes different forms.

Levels of immunoreactive calcitonin (iCT) in human seminal plasma were studied after extraction with immobilized antibodies. Immunoextraction of semen was necessary to abolish the high concentrations of proteolytic enzymes that can interfere in radioimmunoassay (RIA). Since proteolysis occurs already during semen liquefaction, iCT levels and chromatographic properties were compared in extracts from liquefied ejaculates, and samples were the proteolysis was suppressed by immediate dilution of the ejaculates with buffer. The levels in the two groups were 81 +/- 43 pg-equivalents ml-1 seminal plasma (mean +/- SD, n = 6) in liquefied samples and 120 +/- 42 pg-equivalents ml-1 (n = 7) in diluted samples, which is about ten times the iCT levels in extracted blood plasma from healthy males. Extracted liquefied seminal plasma from two vasectomized men contained 93 and 107 pg-equivalents of iCT ml-1 seminal plasma, respectively. Gel chromatography of undiluted semen samples disclosed that iCT was composed of forms larger than monomeric CT, the major part with an approximate molecular weight (MW) of 10,000. Two of the six subjects had a smaller amount of iCT eluting with a MW of about 40,000. In diluted samples, iCT seemed to be more heterogenous with approximate molecular weights ranging from 40,000 to fragments smaller than monomeric CT. In some ejaculates, monomeric CT seemed to be present. Undiluted samples from vasectomized men contained iCT with the same chromatographic pattern as in normal men.

Bone changes after castration in rats. A model for osteoporosis.

Bone changes 6-12 weeks after castration have been studied in 25 female and 27 male middle-aged rats. Castrated female rats gained more weight than their controls, but had decreased bone density and calcium and hydroxyproline content per cm3 bone volume of tibia. Castrated male rats did not differ from controls regarding body weight and the bone parameters. No influence of castration on the mechanical strength of the femora could be detected in either sex. At 2 weeks after castration, the circulating levels of immunoreactive calcitonin (iCT) were decreased in female rats compared to controls. In contrast, iCT was increased both in castrated male and female rats 10 weeks later. We conclude that castration of 6-month-old female rats causes osteoporosis, and therefore represents a promising experimental model for studying postmenopausal bone loss.

Production of immunoreactive calcitonin and parathyroid hormone by embryonal carcinoma cells: Alteration with retinoic acid-induced differentiation

To determine possible ectopic production of, and altered responsiveness to, specific hormones and growth factors which may be involved in mediating embryonic differentiation and development embryonal carcinoma cells in culture have been employed to serve as an in vitro system of embryogenesis. Exposure of F9 embryonal carcinoma cells to all- trans-retinoic acid previously has been shown to induce differentiation of these undifferentiated stem cells to parietal endoderm and to markedly alter the ability of calcitonin and parathyroid hormone to stimulate adenylate cyclase activity. Evidence is presented that F9 cells secrete immunoreactive calcitonin into the culture medium (200 pg/12 hr/10 7 cells) while parietal yolk sac (PYS) cells secrete immunoreactive parathyroid hormone (800 pg/12 hr/10 7 cells). Retinoic-induced differentiation of F9 cells to endoderm results in a progressive reduction in immunoreactive calcitonin production, while there is an increase in the level of immunoreactive parathyroid hormone found in the conditioned medium. After exposure of F9 cells to retinoic acid for 5 days, little calcitonin is detectable in 12-hr conditioned medium. Changes in the intracellular levels of immunoreactive calcitonin and PTH follow a pattern similar to that noted for changes in the amount of secreted hormones. Thus, immunoreactive calcitonin is produced by undifferentiated F9 cells which possess a calcitonin responsive adenylate cyclase system, while parathyroid hormone is produced by parietal endoderm cells which respond to parathyroid hormone with increased cyclic AMP synthesis. Sephadex G50 gel filtration of F9-conditioned medium shows two peaks of immunoreactive calcitonin with M r of 3500 and 20,000. Immunoprecipitation of calcitonin from 35S-labeled F9 cells reveals a specific band of 20,000 M r. Likewise, two peaks of parathyroid hormone immunoreactive material of M r 8000 and 39,000 are noted after gel filtration of PYS cell-conditioned medium, whereas parathyroid hormone immunoprecipitation from the same cells reveals a specific band of 39,000 M r. These results raise the possibility that embryo production of these two hormones at specific stages in development may contribute to the regulation of subsequent steps of differentiation.

Immunoreactive Calcitonin Production by a Human Promyelocytic Leukemia Cell Line HL60

Using a sensitive radioimmunoassay, we detected human immunoreactive calcitonin in cell extracts and in cell-exposed media of the HL60 cell line derived originally from a patient with acute promyelocytic leukemia. The cell extract was chromatographed on a reverse-phase high-pressure liquid chromatography system. Radioimmunoassay of the fractions showed that the immunoreactive calcitonin was heterogeneous but had peaks corresponding to those of synthetic human calcitonin monomer and its sulphoxide. We have previously shown that levels of immunoreactive calcitonin are elevated in the plasma of the majority of patients with acute and chronic myeloid leukemias. Our studies with the HL60 cell line add further support to the concept that leukemic cells can synthesize immunoreactive calcitonin "ectopically."

Immunoreactive calcitonin production by a human promyelocytic leukemia cell line HL60

Using a sensitive radioimmunoassay,we detected human immunoreactive calcitonin in cell extracts and in cell-exposed media of the HL60 cell line derived originally from a patient with acute promyelocytic leukemia. The cell extract was chromatographed on a reverse-phase high- pressure liquid chromatography system. Radioimmunoassay of the fractions showed that the immunoreactive calcitonin was heterogeneous but had peaks corresponding to those of synthetic human calcitonin monomer and its sulphoxide. We have previously shown that levels of immunoreactive calcitonin are elevated in the plasma of the majority of patients with acute and chronic myeloid leukemias. Our studies with the HL60 cell line add further support to the concept that leukemic cells can synthesize immunoreactive calcitonin “ectopically.”

Ectopically produced calcitonin in human hemoblastoses.

The incidence of elevated serum levels of immunoreactive calcitonin (CT) in human myeloproliferative and lymphoproliferative disorders was investigated. On the basis of twice the normal range, about 45% of patients with acute leukemia and blast crisis of chronic myelocytic leukemia (CML) showed elevated serum levels of CT. Markedly elevated levels (greater than 1,000 pg/ml) were only found in this group. Since immunoreactive CT dropped to normal or only slightly elevated levels in remission and increased again before or during relapse, serum CT levels seem to reflect the activity of the disease. However, in patients with chronic leukemia, Hodgkin's and non-Hodgkin's lymphoma, a lower incidence and only slightly elevated serum levels were found. In addition, the molecular weight of the proteohormone in serum specimen and cell extracts was investigated by gel chromatography. Besides physiological CT, different high-molecular weight forms of the hormone could be demonstrated in serum and in cell extracts. Extracts of leukemic cells revealed higher molecular forms only. It is suggested that the proteohormone is ectopically produced by leukemic cells.

Hypocalcemia in infants of diabetic mothers. Studies in circulating calciotropic hormone concentrations.

Twenty-two infants of diabetic mothers (IDM) were studied and were divided into two groups: a first group of 14 IDM did not receive vitamin D3 and was studied at birth and at 2, 24, 48 and 120 hours; a second group was given daily dosage of 60 microgram of vitamin D3 from 3 hours to 120 hours and was studied at 2 hours and 120 hours. In the first group, serum calcium levels decreased markedly during the first 24 hours of life (mean +/- SD: 1.77 +/- 0.3 mmol/l, p less than 0.01) and remained low at 5 days. Serum phosphorus levels remained normal but serum magnesium levels decreased significantly at 24 hours (mean +/- SD: 0.64 +/- 0.108 mmol/l, p less than 0.01) and returned to normal at 5 days. Serum immunoreactive parathormone levels increased consistently to high levels at 24 hours and remained elevated at 120 hours (p less than 0.001). Serum immunoreactive calcitonin levels increased at 24 hours (p less than 0.001) and decreased at 120 hours to low or undetectable values in all infants. In group II, serum 25O-HD levels and 1.25 OH2 D levels increased significantly (p less than 0.001) respectively to 27.2 +/- 2.7 ng/ml and 114 +/- 20 pg/ml at 5 days. The results of this study show hypocalcemia to be a common event in IDM during the first days of life and furthermore hypophosphatemia, hypoparathyroidism, hypomagnesemia or defect of vitamin D metabolism would not seem to be the main etiological factors.

Value of serum calcitonin estimation in clinical oncology.

In 132 consecutive patients with carcinoma of various organs, a higher serum immunoreactive calcitonin (ICT) concentration (median level 50 pg/ml) was found than in 68 normal subjects (median level 20 pg/ml). The incidence of hypercalcitoninaemia was 40%. All 9 patients with primary liver-cell carcinoma were hypercalcitoninaemic. On the other hand, none of the 7 patients with a carcinoma of the breast had raised ICT levels. In bronchogenic cancer a relationship between ICT and cell type was found, with a predominance of high ICT in patients with oatcell and other undifferentiated types, whereas in squamous-cell carcinomas and adenocarcinomas of the lung hypercalcitoninaemia was relatively rare. When we divided all our patients according to differentiation of the tumour cell, it was found that the lower the degree of differentiation, the higher the ICT concentration, whereas opposite results were observed for CEA. When ICT and CEA were estimated concurrently, we found at least one marker increased in 70% of our patients. Our results demonstrate that patients with metastases in the liver have more frequently and increased ICT. In addition, we conclude that lifespan can be expected to be lower in patients with high ICT levels. In a longitudinal study of 46 patients, there was a positive correlation between change in serum ICT and tumour mass.

Heterogeneity of Adult Hypophosphatasia Report of Severe and Mild Cases

Two cases of women with adult hypophosphatasia illustrate the clinical spectrum and potential difficulties in the diagnosis of this condition. Both patients had subnormal serum alkaline phosphatase activity, absence of leukocyte alkaline phosphatase, increased amounts of urinary phosphoethanolamine, and normal levels of immunoreactive calcitonin and parathyroid hormone. In undecalcified bone biopsy specimens, the number of osteoblasts and the tetracycline-labeled calcification front were similar in the two patients, although the percentage of unmineralized bone matrix and the extent of osteoid-covered bone surface were different. Twenty years of bone pain, severe skeletal deformities, and a generalized increase of osteoid in one patient contrasted with an 18-month history of bone pain and patchy osteoid in the other. These cases suggest that adult hypophosphatasia is a heterogeneous disorder and may be more common than previously realized.

Effect of early oral calcium supplementation on serum calcium and immunoreactive calcitonin concentration in preterm infants.

Oral calcium supplements (80 mg/kg per 24 h) were given to 23 preterm infants, and the course of serum calcium, magnesium, immunoreactive calcitonin, and gastrin was compared with a control group of 23 matched infants. In the supplemented group, serum calcium concentrations remained at the baseline level (2.31 mmol/l +/- 0.18 SD) while a fall (from 2.27 +/- 0.18 to 1.91 +/- 0.24 mmol/l) was observed at 12-16 hours of age in the control group, with 4 values < 1.75 mmol/l. There was no change in serum magnesium concentration in either group. The postnatal rise of serum immunoreactive calcitonin concentrations in the control group (from 171 +/- 135 to 493 +/- 273 pg/ml at 12-48 hours of age) was not found in the supplemented group. There was a negative correlation between serum calcium and immunoreactive calcitonin levels in the control group, but not in the supplemented group. There was no correlation between serum immunoreactive calcitonin and gastrin concentrations. These data show that oral calcium supplementation can prevent early neonatal hypocalcaemia, and suggest that this effect is achieved at least in part through a reduction of the postnatal rise of serum immunoreactive calcitonin.

Regulation of calcitonin secretion in normal man by changes of serum calcium within the physiologic range.

To examine the relative importance of calcium and gastrin in regulation of calcitonin secretion, we administered graded oral doses of calcium to 10 normal men, ages 23-29 yr. Each subject had previously shown an appropriate increase in calcitonin secretion in response to a pharmacologic (0.5 mug/kg) pentagastrin injection. On separate days and in random order, each man drank 250 ml of distilled water containing 0.0, 0.5, 1.5, and 3.0 g of elemental calcium as the gluconate salt. Blood samples were drawn before and at 30, 60, 90, 120, 180, and 240 min after the oral calcium dose. The samples were analyzed for calcium by atomic absorption spectroscopy, and for gastrin and calcitonin by radioimmunoassays of established sensitivity and specificity. Ingestion of water (control) caused no change in any of the three variables. Calcium ingestion resulted in dose-related increases, within the normal range, of all three variables. Immunoreactive gastrin rose promptly, peaking at 30 min, and returning to basal levels or below by 120 min. In contrast, calcium and immunoreactive calcitonin levels rose slowly and in parallel, peaking at 120-240 min. Changes in calcitonin and changes in calcium were strongly and positively correlated, r = 0.73, when all data were pooled. Furthermore, individual linear regressions for changes in calcitonin and calcium levels (calculated separately for the three oral calcium doses in each subject) had positive slopes in 28 out of 30 sets (P < 0.01). The changes in calcitonin concentrations were much more poorly correlated with the corresponding changes in serum gastrin levels; in fact, the regression coefficient was weakly negative, r = -0.20. These results show that, at least in young adult men, changes of ambient calcium concentration within the normal range may be of major importance in physiologic regulation of calcitonin secretion. The findings are consistent with the hypothesis that calcitonin functions to prevent excessive postprandial hypercalcemia.

EFFECT OF RAPID VARIATION OF RENAL FUNCTION ON PLASMA CALCITONIN AND PARATHYROID HORMONE IN MAN

Plasma levels of immunoreactive calcitonin (iCT) and parathyroid hormone (iPTH) have been measured sequentially in 6 patients following successful renal transplantation (RT) and in 3 patients during the recovery phase of acute renal failure (ARF). iCT and iPTH returned to normal values within a few days when glomerular filtration improved; both hormones rose in cases of acute graft rejection. Unlike iPTH iCT did not follow closely the variations of creatinine, iCT even rising before creatinine in one graft rejection. These observations could possibly be explained by a dissociation between the renal metabolism of iCT and the glomerular filtration.

ETHANOLINDUCED SECRETION OF CALCITONIN IN CHRONIC RENAL DISEASE

Whisky (25-50 ml) increased plasma levels of immunoreactive calcitonin (iCT) in seventeen of nineteen patients with chronic renal failure. The effect was greater in patients with high levels of iCT than in those with normal levels. Changes in plasma iCT were not related to changes in calcium, phosphate or immunoreactive gastrin, but were inhibited by the prior administration of propranolol.

Hypocalcemia with hypoparathyroidism and renal tubular dysfunction associated with aminoglycoside therapy.

Seventeen patients with malignant disease developed a complex metabolic syndrome of 2-8 weeks' duration characterized by hypocalcemia, hypomagnesemia and hypokalemia following administration of the aminoglycoside group of antibiotics. Gentamicin, Tobramycin, Amikacin, and Sisomicin were all involved. Other features noted were hypoalbuminemia, hypophosphatemia, and hypouricemia. Low immunoreactive parathyroid hormone (i-PTH) levels in the presence of hypocalcemia and absence of hyperplastic changes in the parathyroid gland examined at postmortem confirmed a diagnosis of hypoparathyroidism. Immunoreactive calcitonin levels (i-CT) were not elevated. Renal tubular wasting of potassium and magnesium was documented in six patients and excessive urinary loss of sodium, phosphate, and uric acid was noticed. Twelve patients died before recovering from the metabolic stress and five patients developed progressive renal impairment. A possible potentiating action of chemotherapeutic agents, especially Adriamycin, is suggested.

Plasma calcitonin in Paget's disease of bone.

1. Plasma levels of immunoreactive calcitonin were measured in 22 patients with untreated Paget's disease of bone and in 22 control subjects matched for age and sex. 2. No significant differences in plasma calcitonin were found between patients and control subjects, and hormone levels did not correlate significantly with activity of plasma alkaline phosphatase. 3. These results suggest that Paget's disease of bone is not due to deficient of endogenous calcitonin.